ANTIBODY THERAPY FOR BREAST CANCER: INVESTIGATION OF IMMUNE MODULATION WITH IL-21

在美国2006年将确诊的21.5万例新乳腺癌病例中，约有25% 将过度表达HER2/neu原癌基因。只有大约一半的患者将受益于 人源化抗HER2/neu单抗曲妥珠单抗的应用 化疗没有一种是可以治愈的。曲妥珠单抗治疗的作用机制尚不明确 尽管先天免疫细胞可能携带Fc区(或“恒定”区)的受体(R) 免疫球蛋白也参与其中。我们假设这些携带FCR的细胞与特定的 激活因子可显著增强抗体包被的肿瘤细胞的免疫应答。事实上，我们 发现用白介素21(IL-21)和固定化免疫球蛋白处理自然杀伤(NK)细胞可导致 与细胞相比协同产生干扰素-γ和T细胞趋化因子 单用IL-21或免疫球蛋白治疗。通过IL-21R和FcpR111a以这种方式共同刺激NK细胞导致 丝裂原活化蛋白(MAP)激酶Erk的长时间激活(这对NK细胞至关重要 细胞因子分泌)和AP-1转录因子的协同诱导。我们还发现， 协同刺激的NK细胞协同产生干扰素-K依赖于特定的 NK细胞膜内的信号平台称为脂筏。我们的小鼠研究表明IL-21 显著增强小鼠抗HER2/neu乳腺癌单抗的抗肿瘤活性 作用依赖于内源性产生的干扰素-K。其他细胞因子也被用于 与mAbs的结合不算成功，但IL-21的独特作用使其成为治疗 曲妥珠单抗治疗环境中的应用：IL-21耐受性良好，能够激活NK细胞和CDS* T细胞，并诱导一系列独特的免疫激活细胞因子。在这项提案中，我们将调查 脂筏和ERK诱导的AP-1刺激干扰素-γ基因表达的作用及其机制 干扰素-γ促进接受IL-21和抗HER2单抗的小鼠存活。我们还计划进行一次临床 IL-21联合曲妥珠单抗/紫杉醇治疗局部晚期转移性肿瘤的临床研究 乳腺癌。患者肿瘤和外周血免疫细胞将在治疗前后进行评估 治疗，以确定负责肿瘤根除的机制。临床前和临床 本项目中提出的实验旨在阐明 给予IL-21可增强曲妥珠单抗的活性。这一信息应该会导致进一步的临床 将测试这种治疗组合在HER2(+)乳腺癌患者中的疗效的试验。我们 相信这些研究将确定新的策略，以调节NK细胞的激活，以响应抗体- 包被的靶标，这一信息将导致单抗治疗癌症的改进。

Of the 215,000 new cases of breast cancer that will be diagnosed in the United States in 2006, about 25% will over-express the HER2/neu proto-oncogene. Only about half of these patients will benefit from administration of the humanized anti-HER2/neu monoclonal antibody (mAb)trastuzumab with chemotherapy and none will be cured. There is no clear mechanism of action for trastuzumab therapy although it is possible that innate immune cells bearing receptors (R) for the Fc (or "constant") region of immunoglobulin are involved. We hypothesized that co-stimulation of these FcR-bearing cells with specific activating factors would significantly enhance the immune response to Ab-coated tumor cells. Indeed, we found that treatment of natural killer (NK) cells with interleukin-21 (IL-21) and immobilized IgG led to synergistic production of interferon-gamma (IFN-K) and T cell-attracting chemokines as compared to cells treated with IL-21 or IgG alone. Co-stimulation of NK cells in this manner via the IL-21R and FcpRllla led to prolonged activation of the mitogen-activated protein (MAP) kinase Erk (which was critical for NK cell cytokine secretion) and synergistic induction of the AP-1 transcription factor. We also found that the synergistic production of IFN-K by co-stimulated NK cells was dependent upon the presence of specialized signaling platforms within the NK cell membrane called lipid rafts. Our murine studies showed that IL-21 significantly enhanced the anti-tumor activity of a murine anti-HER2/neu breast cancer mAb and that this effect was dependent upon endogenously-produced IFN-K. Other cytokines have been employed in combination with mAbs with modest success, but the unique actions of IL-21 make it a superior choice for use in the setting of trastuzumab therapy: IL-21 is well-tolerated, is able to activate both NK cells and CDS* T cells, and induces a unique array of immune activating cytokines. In this proposal we will investigate the role of lipid rafts and Erk-induced AP-1 in stimulating IFN- y gene expression and the mechanism by which IFN-y promotes survival in mice receiving IL-21 and the anti-HER2 mAb. We also plan to conduct a clinical trial of IL-21 in combination with trastuzumab/paclitaxel in patients with metastatic and locally-advanced breast cancers. Patient tumors and peripheral blood immune cells will be evaluated before and after treatment in order to identify the mechanisms responsible for tumor eradication. The pre-clinical and clinical experiments proposed in this project are designed to elucidate the specific mechanisms by which administration of IL-21 enhances the activity of trastuzumab. This information should lead to further clinical trials that will test the efficacy of this therapeutic combination in patients with HER2 (+) breast cancer. We believe that these studies will identify new strategies for modulating NK cell activation in response to Ab- coated targets and that this information will lead to improvements in the mAb therapy of cancer.